TCR-mimic bispecific antibodies targeting LMP2A show potent activity against EBV malignancies
Mahiuddin Ahmed, Andres Lopez-Albaitero, Dmitry Pankov, Brian H. Santich, Hong Liu, Su Yan, Jingyi Xiang, Pei Wang, Aisha N. Hasan, Annamalai Selvakumar, Richard J. O’Reilly, Cheng Liu, Nai-Kong V. Cheung
Mahiuddin Ahmed, Andres Lopez-Albaitero, Dmitry Pankov, Brian H. Santich, Hong Liu, Su Yan, Jingyi Xiang, Pei Wang, Aisha N. Hasan, Annamalai Selvakumar, Richard J. O’Reilly, Cheng Liu, Nai-Kong V. Cheung
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Research Article Oncology Therapeutics

TCR-mimic bispecific antibodies targeting LMP2A show potent activity against EBV malignancies

Abstract

EBV infection is associated with a number of malignancies of clinical unmet need, including Hodgkin lymphoma, nasopharyngeal carcinoma, gastric cancer, and posttransplant lymphoproliferative disease (PTLD), all of which express the EBV protein latent membrane protein 2A (LMP2A), an antigen that is difficult to target by conventional antibody approaches. To overcome this, we utilized phage display technology and a structure-guided selection strategy to generate human T cell receptor–like (TCR-like) monoclonal antibodies with exquisite specificity for the LMP2A-derived nonamer peptide, C426LGGLLTMV434 (CLG), as presented on HLA-A*02:01. Our lead construct, clone 38, closely mimics the native binding mode of a TCR, recognizing residues at position P3–P8 of the CLG peptide. To enhance antitumor potency, we constructed dimeric T cell engaging bispecific antibodies (DiBsAb) of clone 38 and an affinity-matured version clone 38-2. Both DiBsAb showed potent antitumor properties in vitro and in immunodeficient mice implanted with EBV transformed B lymphoblastoid cell lines and human T cell effectors. Clone 38 DiBsAb showed a stronger safety profile compared with its affinity-matured variant, with no activity against EBV– tumor cell lines and a panel of normal tissues, and was less cross-reactive against HLA-A*02:01 cells pulsed with a panel of CLG-like peptides predicted from a proteomic analysis. Clone 38 was also shown to recognize the CLG peptide on other HLA-A*02 suballeles, including HLA-A*02:02, HLA-A*02:04, and HLA-A*02:06, allowing for its potential use in additional populations. Clone 38 DiBsAb is a lead candidate to treat EBV malignancies with one of the strongest safety profiles documented for TCR-like mAbs.

Authors

Mahiuddin Ahmed, Andres Lopez-Albaitero, Dmitry Pankov, Brian H. Santich, Hong Liu, Su Yan, Jingyi Xiang, Pei Wang, Aisha N. Hasan, Annamalai Selvakumar, Richard J. O’Reilly, Cheng Liu, Nai-Kong V. Cheung

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Figure 5

DiBsAbs 38 and 38-2 show potent T cell dependent cytotoxicity of EBV+HLA-A*02:01+ tumor cell targets but not EBV– or HLA-A*02:01– targets.

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DiBsAbs 38 and 38-2 show potent T cell dependent cytotoxicity of EBV+HLA...
Cytotoxicity and specificity of clone 38 (blue squares) and 38-2 (orange triangles) and control (black circles) DiBsAb was measured against 8 cell lines. Activated cord blood T cells were incubated with each cell line for 4 hours (10:1 E:T). Each curve represents the cytotoxicity of a serial titration of each DiBsAb against HLA-A*02:01+EBV+ tumor cell lines (top row), and target negative cell lines (bottom row). HLA-A*02:01+EBV+ cell lines: RPMI-6666, DT BLCL, F BLCL, and HONE-1-A2. Target negative cell lines: K BLCL [HLA-A*02:01–EBV+], MCF7, COLO205, and HEPG2 [HLA-A*02:01+EBV–]. Data are from 3 technical replicates per experimental condition with mean ± SD plotted. EC50 values were determined using a nonlinear fitting algorithm (log [agonist] vs. response-variable slope with 4 parameters) and are shown in Supplemental Table 5.